Pneumocystis entered the spotlight of public health as the hallmark of HIV infection. As research on the fungus expanded, Pneumocystis pneumonia was found to infect a variety of patients with the commonality of a suppressed immune system. As the length of survival in AIDS, cancer, transplant, and other immunosuppressed patients has been prolonged, the prevalence rate of Pneumocystis pneumonia in the population also has increased. Treatments for Pneumocystis pneumonia in such individuals often cause adverse side effects and are not always effective. Therefore, developing an effective vaccine is of great interest for researchers and the medical community.
Pneumocystis cannot be continuously cultured outside of its host. Pneumocystis also has a host species-dependent specificity which complicates the ability to use animal derived organisms to immunize humans. Pneumocystis organisms derived from different hosts have isoform variants of common antigens resulting in different (i.e., non-crossreactive) antigenic determinants (Gigliotti et al., “Antigenic Characterization of Pneumocystis carinii,” Semin. Respir. Infect. 13:313-322 (1998); Gigliotti et al., “Further Evidence of Host Species-Specific Variation in Antigens of Pneumocystis carinii Using the Polymerase Chain Reaction,” J. Infect. Dis. 168:191-194 (1993)). Attempts to infect laboratory animals with Pneumocystis isolated from heterologous mammalian species have met with little to no success (Aliouat et al., “Pneumocystis Cross Infection Experiments Using SCID Mice and Nude Rats as Recipient Host, Showed Strong Host-Species Specificity,” J. Eukaryot. Microbiol. 41:71S (1994); Atzori et al., “P. carinii Host Specificity: Attempt of Cross Infections With Human Derived Strains in Rats,” J. Eukaryot. Microbiol. 46:112S (1999); Gigliotti et al., “Pneumocystis carinii Host Origin Defines the Antibody Specificity and Protective Response Induced by Immunization,” J. Infect. Dis. 176:1322-1326 (1997)). However, immunocompetent mice immunized with whole mouse Pneumocystis are protected from developing Pneumocystis pneumonia after T cell depletion and subsequent challenge, whereas unimmunized cohorts are not protected (Harmsen et al., “Active Immunity to Pneumocystis carinii Reinfection in T-cell-depleted Mice,” Infect. Immun. 63:2391-2395 (1995)).
The surface glycoprotein gpA is an abundant and immunodominant antigen of Pneumocystis (Graves et al., “Development and Characterization of Monoclonal Antibodies to Pneumocystis carinii,” Infect. Immun. 51:125-133 (1986)), although immunization with this antigen does not adequately protect against infection in a mouse model of Pneumocystis pneumonia (Gigliotti et al., “Immunization with Pneumocystis carinii gpA is Immunogenic But Not Protective in a Mouse Model of P. carinii Pneumonia,” Infect. Immun. 66:3179-3182 (1998)). The majority of monoclonal antibodies (“mAb”) against Pneumocystis surface antigens react with only isoforms showing host species-specificity identical to that of the immunogen (Gigliotti et al., “Pneumocystis carinii Host Origin Defines the Antibody Specificity and Protective Response Induced by Immunization,” J. Infect. Dis. 176:1322-1326 (1997)). mAb4F11 was obtained by selective screening of anti-mouse Pneumocystis hybridomas for recognition of Pneumocystis antigens other than gpA (Lee et al., “Molecular Characterization of KEX1, a Kexin-Like Protease in Mouse Pneumocystis carinii,” Gene 242:141-150 (2000)). mAb4F11 confers passive prophylaxis against development of Pneumocystis pneumonia when administered intranasally to SCID mice (Gigliotti et al., “Passive Intranasal Monoclonal Antibody Prophylaxis Against Murine Pneumocystis carinii Pneumonia,” Infect. Immun. 70:1069-1074 (2002)). Furthermore, mAb4F11 recognizes surface antigens of Pneumocystis derived from different hosts, including humans. A screen of a Pneumocystis cDNA expression library using mAb4F11 revealed a number of positive clones, including mouse Pneumocystis Kex1 (Lee et al., “Molecular Characterization of KEX1, a Kexin-Like Protease in Mouse Pneumocystis carinii,” Gene 242:141-150 (2000)). Based on sequence homology to its ortholog in Saccharomyces cerevisiae, Kex1 is a member of the kexin family of subtilisin-like proteases (Lee et al., “Molecular Characterization of KEX1, a Kexin-Like Protease in Mouse Pneumocystis carinii,” Gene 242:141-150 (2000)).
CD4 depletion models in mice are designed to mimic individuals with a suppressed adaptive immune response. Injecting with whole Pneumocystis has been shown to provide sterilizing immunity in such immunocompromised mice. However, due to a variety of factors, whole Pneumocystis is not a viable vaccine option in humans. Extensive studies of antibodies to Pneumocystis led to the discovery of the monoclonal antibody 4F11, which is of great scientific relevance because it provides protection against the development of Pneumocystis pneumonia via passive prophylaxis and cross reacts with human-derived Pneumocystis. 4F11 recognizes an epitope that is present on two distinct antigens in mouse-derived Pneumocystis: Kexin and A12. Further analysis showed the 4F11 epitope is on the C-terminal half of the A12 gene. This makes the A12 antigen a candidate for a potential vaccine.
However, attempts to produce a full-length A12 protein in large quantities in yeast and E. coli have been unsuccessful thus far due to problematic codons at the N-terminus.
The present invention is directed to overcoming these and other deficiencies in the art.